Performance characteristics of Fungitell STAT™, a rapid (1→3)-ß-D-glucan single patient sample in vitro diagnostic assay.

Serum (1→3)-ß-D-glucan (BDG), is an adjunct test in the diagnosis of invasive fungal disease (IFD). Fungitell STAT™, a facile, rapid, single patient option, executable for one or more patient specimens in approximately an hour, has been developed to address a need for rapid in-house testing. This method presents qualitative information concerning serum BDG levels, using an index value that allows the rapid categorization of patients as positive, negative, or indeterminate relative to serum BDG titer. The categorical and analytical performance of Fungitell STAT was evaluated. The categorical agreement between methods was established by testing patient samples which had been previously categorized with Fungitell. Receiver Operating Characteristic curves were used to identify cut-offs using 93 de-identified patient specimens. Subsequently, using these cutoffs, an independent group of 488 patient specimens was analyzed. Positive percent agreement (PPA) with, and without, indeterminate results was 74% and 99%, respectively. Negative percent agreement (NPA) was 91% and 98% with, and without, indeterminate results, respectively. Additionally, commercially available normal off-the-clot sera were spiked with Saccharomyces cerevisiae-derived (1→3)-ß-D-glucan to produce analytical samples. Analytical reproducibility using spiked samples was excellent with 94% of the CV (coefficient of variation) values ≤10% among three independent laboratories. Good correlation with the predicate method was demonstrated with correlation coefficients of 0.90 or better with patient samples and 0.99 with spiked samples. The Fungitell STAT index assay provides a rapid and suitable method for serum BDG testing.

Isolation and Identification of Mesenchymal Stem Cells Derived from Adipose Tissue of Sprague Dawley Rats.

Adult mesenchymal cells have revolutionized molecular and cell biology in recent decades. They can differentiate into different specialized cell types, in addition to their great capacity for self-renewal, migration, and proliferation. Adipose tissue is one of the least invasive and most accessible sources of mesenchymal cells. It has also been reported to have higher yields compared to other sources, as well as superior immunomodulatory properties. Recently, different procedures for obtaining adult mesenchymal cells from different tissue sources and animal species have been published. After evaluating the criteria of some authors, we standardized a methodology applicable to different purposes and easily reproducible. A pool of stromal vascular fraction (SVF) from perirenal and epididymal adipose tissue allowed us to develop primary cultures with optimal morphology and functionality. The cells were observed adhered to the plastic surface for 24 h, and exhibited a fibroblast-like morphology, with prolongations and a tendency to form colonies. Flow cytometry (FC) and immunofluorescence (IF) techniques were used to assess the expression of the membrane markers CD105, CD9, CD63, CD31, and CD34. The ability of adipose-derived stem cells (ASCs) to differentiate into the adipogenic lineage was also assessed using a cocktail of factors (4 µM insulin, 0.5 mM 3-methyl-iso-butyl-xanthine, and 1 µM dexamethasone). After 48 h, a gradual loss of fibroblastoid morphology was observed, and at 12 days, the presence of lipid droplets positive to oil red staining was confirmed. In summary, a procedure is proposed to obtain optimal and functional ASC cultures for application in regenerative medicine.